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CD146 Recombinant Rabbit Monoclonal Antibody [JB42-35]

Usd: 385

Specification

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Catalog# ET7107-91

CD146 Recombinant Rabbit Monoclonal Antibody [JB42-35]

Application

  • WB

  • IF-Cell

  • IHC-P

  • FC

Reactivity

  • Human

  • Mouse

Predicted reactivity

  • Rat

Predicted species support after-sales service
Conjugation

  • unconjugated

This product has been cited in peer reviewed publications, see list HERE

Overview

Product Name

CD146 Recombinant Rabbit Monoclonal Antibody [JB42-35]

Antibody Type

Recombinant Rabbit monoclonal Antibody

Immunogen

Synthetic peptide within C-terminal human CD146 (Cytoplasmic).

Species Reactivity

Human, Mouse (Predicted: Rat)

Predicted species support after-sales service

Validated Applications

WB, IF-Cell, IHC-P, FC

Molecular Weight

Predicted band size: 72 kDa

Positive Control

A375 cell lysate, HeLa cell lysate, HUVEC cell lysate, HepG2 cell lysate, Mouse lung tissue lysate, A375, human tonsil tissue.

Conjugation

unconjugated

Clone Number

JB42-35

RRID

AB_3070796

Product Features

Form

Liquid

Concentration

Storage Instructions

Shipped at 4℃. Store at +4℃ short term (1-2 weeks). It is recommended to aliquot into single-use upon delivery. Store at -20℃ long term.

Storage Buffer

1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.

Isotype

IgG

Purification Method

Protein A affinity purified.

Application Dilution

  • WB

  • 1:1,000

  • IF-Cell

  • 1:100

  • IHC-P

  • 1:200

  • FC

  • 1:1,000

Target

Function

The tumorigenic and metastatic phenotype of melanoma cells correlates well with an increased expression of cell-cell and cell-matrix adhesion receptors. The human Mel-CAM gene encodes a transmembrane glycoprotein, also designated MCAM, MUC18 or CD146, that belongs to the immunoglobulin superfamily and functions as a Ca2+-independent cell adhesion molecule. The deduced human sequence of 603 amino acids consists of a signal peptide, five immunoglobulin-like domains, a transmembrane region and a short cytoplasmic tail. Mel-CAM expression is restricted to advanced primary and metastatic melanomas and to cell lines of the neuroectodermal lineage, but not normal melanocytes. Mel-CAM is found on 80% of advanced primary human mela-nomas and correlates well with development of metastatic disease. Mel-CAM activation initiates an outside-in signaling pathway that involves the protein tyrosine kinases Fyn, FAK and paxillin. Mel-CAM influences the dynamics of Actin cytoskeleton rearrangement and is essential for the maintenance of thymic architecture and function.

Background References

1. Johnson J P et al. The progression associated antigen MUC18: a unique member of the immunoglobulin supergene family. Melanoma Res 3:337-340 (1993).

2. Anfosso F et al. Outside-in signaling pathway linked to CD146 engagement in human endothelial cells. J Biol Chem 276:1564-1569 (2001).

Tissue Specificity

Detected in endothelial cells in vascular tissue throughout the body. May appear at the surface of neural crest cells during their embryonic migration. Appears to be limited to vascular smooth muscle in normal adult tissues. Associated with tumor progression and the development of metastasis in human malignant melanoma. Expressed most strongly on metastatic lesions and advanced primary tumors and is only rarely detected in benign melanocytic nevi and thin primary melanomas with a low probability of metastasis.

Subcellular Location

Membrane.

UNIPROT

SwissProt: P43121 Human

SwissProt: Q8R2Y2 Mouse

SwissProt: Q9EPF2 Rat

Synonyms

A32 antigen antibody

CD 146 antibody

CD146 antibody

CD146 antigen antibody

Cell surface glycoprotein MUC18 antibody

Cell surface glycoprotein P1H12 antibody

Gicerin antibody

Mcam antibody

Melanoma adhesion molecule antibody

Melanoma associated antigen A32 antibody

Expand

Images

  • Western blot analysis of CD146 on different lysates with Rabbit anti-CD146 antibody (<a href="/products/ET7107-91" style="font-weight: bold;text-decoration: underline;">ET7107-91</a>) at 1/1,000 dilution.<br /><br />Lane 1: A375 cell lysate (20 µg/Lane)<br />Lane 2: HeLa cell lysate (20 µg/Lane)<br />Lane 3: HUVEC cell lysate (20 µg/Lane)<br />Lane 4: HepG2 cell lysate (20 µg/Lane)<br />Lane 5: Mouse lung tissue lysate (40 µg/Lane)<br /><br />Predicted band size: 72 kDa<br />Observed band size: 120 kDa<br /><br />Exposure time: 14 seconds; ECL: K1801;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET7107-91" style="font-weight: bold;text-decoration: underline;">ET7107-91</a>) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    Western blot analysis of CD146 on different lysates with Rabbit anti-CD146 antibody (ET7107-91) at 1/1,000 dilution.

    Lane 1: A375 cell lysate (20 µg/Lane)
    Lane 2: HeLa cell lysate (20 µg/Lane)
    Lane 3: HUVEC cell lysate (20 µg/Lane)
    Lane 4: HepG2 cell lysate (20 µg/Lane)
    Lane 5: Mouse lung tissue lysate (40 µg/Lane)

    Predicted band size: 72 kDa
    Observed band size: 120 kDa

    Exposure time: 14 seconds; ECL: K1801;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET7107-91) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • <span style="font-weight: bold;">☑ Knockdown (KD)</span><br /><br />Western blot analysis of CD146 on different lysates with Rabbit anti-CD146 antibody (<a href="/products/ET7107-91" style="font-weight: bold;text-decoration: underline;">ET7107-91</a>) at 1/1,000 dilution.<br /><br />Lane 1: HAP1-parental cell lysate<br />Lane 2: HAP1-CD146 KD cell lysate<br /><br />Lysates/proteins at 10 µg/Lane.<br /><br />Predicted band size: 72 kDa<br />Observed band size: 120 kDa<br /><br />Exposure time: 180 seconds; ECL: K1801;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET7107-91" style="font-weight: bold;text-decoration: underline;">ET7107-91</a>) at 1/1,000 dilution was used in K1803 at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    ☑ Knockdown (KD)

    Western blot analysis of CD146 on different lysates with Rabbit anti-CD146 antibody (ET7107-91) at 1/1,000 dilution.

    Lane 1: HAP1-parental cell lysate
    Lane 2: HAP1-CD146 KD cell lysate

    Lysates/proteins at 10 µg/Lane.

    Predicted band size: 72 kDa
    Observed band size: 120 kDa

    Exposure time: 180 seconds; ECL: K1801;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET7107-91) at 1/1,000 dilution was used in K1803 at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • Immunocytochemistry analysis of A375 cells labeling CD146 with Rabbit anti-CD146 antibody (<a href="/products/ET7107-91" style="font-weight: bold;text-decoration: underline;">ET7107-91</a>) at 1/100 dilution.<br /><br />Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-CD146 antibody (<a href="/products/ET7107-91" style="font-weight: bold;text-decoration: underline;">ET7107-91</a>) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor&trade; 488, <a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.<br /><br />Beta tubulin (<a href="/products/HA601187" style="font-weight: bold;text-decoration: underline;">HA601187</a>, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor&trade; 594, <a href="/products/HA1126" style="font-weight: bold;text-decoration: underline;">HA1126</a>) was used as the secondary antibody at 1/1,000 dilution.

    Immunocytochemistry analysis of A375 cells labeling CD146 with Rabbit anti-CD146 antibody (ET7107-91) at 1/100 dilution.

    Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-CD146 antibody (ET7107-91) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

    Beta tubulin (HA601187, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

  • Immunohistochemical analysis of paraffin-embedded human tonsil tissue with Rabbit anti-CD146 antibody (<a href="/products/ET7107-91" style="font-weight: bold;text-decoration: underline;">ET7107-91</a>) at 1/200 dilution.<br /><br />The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET7107-91" style="font-weight: bold;text-decoration: underline;">ET7107-91</a>) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded human tonsil tissue with Rabbit anti-CD146 antibody (ET7107-91) at 1/200 dilution.

    The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7107-91) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Immunohistochemical analysis of paraffin-embedded human tonsil tissue with Rabbit anti-CD146 antibody (<a href="/products/ET7107-91" style="font-weight: bold;text-decoration: underline;">ET7107-91</a>) at 1/200 dilution.<br /><br />The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET7107-91" style="font-weight: bold;text-decoration: underline;">ET7107-91</a>) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded human tonsil tissue with Rabbit anti-CD146 antibody (ET7107-91) at 1/200 dilution.

    The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7107-91) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Flow cytometric analysis of A375 cells labeling CD146.<br /><br />Cells were fixed and permeabilized. Then stained with the primary antibody (<a href="/products/ET7107-91" style="font-weight: bold;text-decoration: underline;">ET7107-91</a>, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor&trade; 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (<a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

    Flow cytometric analysis of A375 cells labeling CD146.

    Cells were fixed and permeabilized. Then stained with the primary antibody (ET7107-91, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

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Citation

Products with the same target and pathway

metafield image

CD146 Rabbit Polyclonal Antibody

Application: WB,IP

Reactivity: Human,Mouse,Rat

Conjugate: unconjugated